Cytotoxic T lymphocytes (CTLs) may play a role in host defence against hepatitis C virus (HCV) infection, and HCV-specific CTL epitopes may be included in vaccines to induce protective CTLs. We identified three new epitopes within the HCV nucleoprotein recognized by CTLs. HCV nucleoprotein residues 28-37 are the minimal epitope recognized by CTLs in association with the class I human leukocyte antigen B60, and epitopes in HCV nucleoprotein residues 111-130 and 161-180 are both recognized by CTLs in association with the class II human leukocyte antigen DRBI*08032.